We investigated the effects of PI(3,4)P2 and PI(3,4,5)P3 on the activities of PLC isozymes. PI(3,4,5)P3 increased the activities of PLC- gamma1 and PLC-gamma2 but had no effect on PLC-beta1 or PLC- delta. PI(3,4,5)P3 stimulated the activities of PLC- gamma1 and PLC-gamma2 by 8- fold, whereas PI(3,4)P2 had no effect on the activities of any of the PLC isozymes examined. We prepared various recombinant proteins containing different domains of PLC-gamma1and measured their effects on the activity of PLC- gamma1. All SH2 domain, containing proteins inhibited the PI(3,4,5)P3-stimulated activity of PLC-gamma1 in a concentration, dependent manner, whereas proteins corresponding to the SH3 and PH domains had no effect. These results suggest that PI(3,4,5)P3 activates PLCgamma isozymes by binding to their SH2 domains. To determine whether activation of PLC by PI(3,4,5)P3 could be detected in intact cells, we transiently expressed in COS-7 cells the 110-kD subunit (p110) of PI 3-kinase with farnesylation signal (CAAX) sequences and measured the release of inositol phosphates. Expression of p110- CAAX induced a 45% increase in the amount of inositol phosphates, and this effect was blocked by pretreatment of cells with LY294002, a specific inhibitor of PI 3-kinase. These results suggest that PI(3,4,5)P3 generated by the activated p110 subunit was able to activate PLC. We also studied the effect of LY294002 on PI(4,5)P2 hydrolysis induced by PDGF in NIH 3T3 cells. Whereas PDGF induced an 8- fold increase in PLC activity in control cells, pretreatment of cells with LY294002 reduced this response by 40%. The presence of the inhibitor affected neither the basal amount of inositol phosphates nor the tyrosine phosphorylation of PLC- gamma1 and PDGF receptor.These results suggest that receptors coupled to PI 3- kinase may activate PLC- gamma isozymes indirectly, in the absence of PLC- gamma tyrosine phosphorylation, through the generation of PI(3,4,5)P3.